| HO-1在巨噬细胞中的抗炎和抗氧化作用 |
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| 引用本文： | 付京城,何俊辉,仝江,吴魏,郭爽,杨彦宾,韩莹倩,王月影,李和平. HO-1在巨噬细胞中的抗炎和抗氧化作用[J]. 中国畜牧兽医, 2023, 50(2): 807-816. DOI: 10.16431/j.cnki.1671-7236.2023.02.038 |
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| 作者姓名： | 付京城 何俊辉 仝江 吴魏 郭爽 杨彦宾 韩莹倩 王月影 李和平 |
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| 作者单位： | 1. 河南农业大学, 农业农村部动物生化与营养重点实验室, 郑州 450046;2. 河南省动物生长发育调控重点实验室, 郑州 450046 |
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| 基金项目： | 河南省科技攻关计划(212102110356、222102110402) |
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| 摘 要： | 【目的】旨在揭示血红素加氧酶1(HO-1)在巨噬细胞中的抗炎和抗氧化作用。【方法】利用不同浓度脂多糖(LPS,0、3、5、10、15、20、25μg/mL)、HO-1(0、0.02、0.04、0.06、0.08、0.10μg/mL)及锌原卟啉(zinc protoporphyrin, ZPP;0、5、10、15、20、30 ng/mL)分别处理巨噬细胞(RAW264.7),12 h后通过CCK8法检测RAW264.7细胞活力，计算LPS、HO-1和ZPP处理RAW264.7细胞的最佳浓度。将RAW264.7细胞随机分为对照组(CT)、LPS组(LPS)、LPS+HO-1组(LH)、HO-1组(HO-1)、LPS+HO-1+ZPP组(LHZ),每组3个重复。CT组细胞用含10%胎中血清的DMEM培养基培养；LPS组细胞用LPS处理；LH组细胞用LPS和HO-1共处理；HO-1组细胞用HO-1处理；LHZ组细胞用LPS、HO-1和ZPP共处理，各组细胞的处理时间均为12 h,收集细胞和上清。采用ELISA法检测上清液中白细胞介素-6(IL-6)、IL-8、肿瘤坏死因子α(TNF-α)、活性...
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| 关 键 词： | 血红素加氧酶-1 脂多糖 RAW264.7细胞 炎症反应 氧化应激 |
| 收稿时间： | 2022-07-13 |
Anti-inflammatory and Antioxidant Roles of HO-1 in Macrophages |
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| FU Jingcheng,HE Junhui,TONG Jiang,WU Wei,GUO Shuang,YANG Yanbin,HAN Yingqian,WANG Yueying,LI Heping. Anti-inflammatory and Antioxidant Roles of HO-1 in Macrophages[J]. China Animal Husbandry & Veterinary Medicine, 2023, 50(2): 807-816. DOI: 10.16431/j.cnki.1671-7236.2023.02.038 |
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| Authors: | FU Jingcheng HE Junhui TONG Jiang WU Wei GUO Shuang YANG Yanbin HAN Yingqian WANG Yueying LI Heping |
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| Affiliation: | 1. Key Laboratory of Animal Biochemistry and Nutrition, Ministry of Agriculture and Rural Affairs, Henan Agricultural University, Zhengzhou 450046, China;2. Key Laboratory of Animal Growth and Development of Henan Province, Zhengzhou 450046, China |
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| Abstract: | 【Objective】 This study was aim to reveal the anti-inflammatory and antioxidant effects of heme oxygenase-1 (HO-1) in macrophages.【Method】 Different concentrations of lipopolysaccharide (LPS,0,3,5,10,15,20,25 μg/mL),HO-1(0,0.02,0.04,0.06,0.08,0.10 μg/mL) and zinc protoporphyrin (ZPP;0,5,10,15,20,30 ng/mL) respectively were used to treat macrophages (RAW264.7) for 12 hours.The viability of RAW264.7 cells was detected by CCK8 method,and the optimal concentration of RAW264.7 cells treated with LPS,HO-1 and ZPP was calculated. RAW264.7 cells were randomly divided into control group (CT),LPS group (LPS),LPS+HO-1 group (LH),HO-1 group (HO-1),LPS+HO-1+ ZPP group (LHZ),with three replicates in each group.The cells in CT group were seeded in DMEM medium containing 10% fetal bovine serum.The cells in LPS group were treated with LPS.The cells in LH group were treated with LPS and HO-1.The cells in HO-1 group were treated with HO-1.The cells in LHZ group were co-cultured with LPS,HO-1 and ZPP.Cells in each group was treated for 12 hours,collecting the cells and supernatant.The contents of interleukin-6 (IL-6),IL-8,tumor necrosis factor-α (TNF-α), reactive oxygen sepecies (ROS), malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GPx) in the supernatant were detected by ELISA.The mRNA expression of IL-6,IL-8,TNF-α,nuclear factor-E2-related factor 2 (Nrf2) and HO-1 was measured by quantitative Real-time PCR.The expression of Nrf2,HO-1 protein was detected by Western blotting.【Result】 Compared with cells treated by 0 μg/mL LPS,the concentration of 20 and 25 μg/mL LPS extremely significantly decreased the viability of RAW264.7 cells (P＜0.01).Compared with cells treated by 0 μg/mL HO-1,the concentration of 0.08 and 0.10 μg/mL HO-1 extremely significantly increased cell viability (P＜0.01).Compared with cells treated by 0 ng/mL,ZPP with 15,20 and 30 ng/mL extremely significantly reduced cell viability (P＜0.01).The results showed that the concentration of LPS,HO-1 and ZPP treatment was 20 μg/mL,0.08 μg/mL,15 ng/mL in subsequent experiment,respectively.Compared with CT group,IL-6,IL-8 and TNF-α mRNA expression in LPS group were extremely significantly up-regulated (P＜0.01).The contents of IL-6,IL-8,TNF-α,MDA and ROS were extremely significantly increased (P＜0.01),and the contents of GPx and SOD were extremely significantly decreased (P＜0.01).Compared with LPS group,IL-6,IL-8 and TNF-α mRNA expression in LH group were extremely significantly decreased (P＜0.01).The contents of IL-6,IL-8,TNF-α,MDA and ROS were extremely significantly decreased (P＜0.01),and the contents of GPx and SOD were extremely significantly increased (P＜0.01).Compared with the LH group,IL-6,IL-8 and TNF-α mRNA expression in LHZ group were extremely significantly up-regulated (P＜0.01).The content of IL-6,IL-8,TNF-α,MDA and ROS were extremely significantly increased (P＜0.01),while the contents of GPx and SOD were extremely significantly decreased (P＜0.01).The results of Nrf2/HO-1 signal pathway molecules expression showed that the mRNA and protein expressions of Nrf2 and HO-1 in LPS group were extremely significantly up-regulated compared with CT group (P＜0.01).Compared with LPS group,Nrf2 and HO-1 mRNA expression in LH group were extremely significantly down-regulated (P＜0.01),Nrf2 protein expression was extremely significantly decreased (P<0.01),and HO-1 protein expression was extremely significantly increased (P＜0.01).Compared with LH group,Nrf2 and HO-1 mRNA expression in LHZ group was extremely significantly up-regulated (P＜0.01),Nrf2 protein expression was extremely significantly increased (P＜0.01),and HO-1 protein expression was extremely significantly decreased (P＜0.01).Compared with CT group,the indexes of HO-1 group were not significantly different (P>0.05).【Conclusion】 20 μg/mL LPS induced macrophages to produce inflammatory response and oxidative stress.0.08 μg/mL HO-1 had anti-inflammatory and antioxidant effects.HO-1 regulated Nrf2 signaling pathway to exert anti-inflammatory and antioxidant effects. |
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| Keywords: | heme oxygenase-1 lipopolysaccharide RAW264.7 cells oxidative stress inflammatory response |
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